X-RAY CRYSTALLOGRAPHY OF MACROMOLECULES

这项建议是对最近在 结构生物学领域。遗传学和生物化学研究进展 产生了对大量的 大分子。X射线仪器和计算硬件的进展 让这一切成为可能。根据我们在开发广泛使用的 大分子X射线结晶学HKL数据还原程序包(Denzo， Scalepack，XdisplayF)我们建议进一步开发这一领域。这个 这项建议的具体目标是： 1)开发不完美晶体的数据简化方法-高度 马赛克的或孪生的。将有可能索引一个以上的衍射 一个探测器图像中的图案，计算衍射之间的重叠 布拉格峰值反射尾部侵入的模式及其校正 彼此之间。 2)将多波长反常色散(MAD)方法扩展到 当前信错比不足以满足 结构确定。这将通过以下方式实现：校准和 对仪器响应系数的修正现在将MAD方法限制为 具有较高反常衍射率的高衍射率晶体 信号。 3)通过提前模拟提供最优的数据收集策略 数据收集预期的倒数空间覆盖率和探测器 作为可能设置的函数的分辨率-数据收集角度和 晶体到探测器的距离。 4)通过在线减少数据和计算来提供实时反馈 综合统计。这对用户来说特别重要 同步加速器设施需要等待数月才能重复其 实验。 5)扩大结晶学实验研究的范围 通过部分重叠的去卷积最优地进行数据简化 反思和三维轮廓拟合，发展自动化和互动式 编辑工具，以最大限度地减少可能的实验影响 减少的数据上的伪像。 6)扩展方法，直观查看数据采集过程 以及将数据从当前原始数据的直接可视化简化为 影响数据质量的因素的综合视觉表示法.平均值 反射剖面、去卷积积分、晶体滑移、探测器 响应函数等。 7)为实验者提供数据库式的组织工具 跟踪整个结晶学项目。该数据库将用于 创建统一的报告-合并统计数据、覆盖范围、衰减、滑动等。 最后，上述所有发展将整合为一个 高效、易用的计算机和探测器通用软件包。 由此产生的软件将提高3D的精度和可靠性 通过x射线结晶学研究大分子的结构，能够解决 晶体结构太难用目前的方法解决，以及 将加快、简化和减少所需的时间和工作量 结构确定过程，加快对生物学的理解 基于系统和结构的药物设计。

This proposal is in response to the recent, dramatic expansion in the fields of structural biology. Progress in genetics and biochemistry created a need for precise stereochemical understanding of large numbers of macromolecules. Progress in X-ray instrumentation and computing hardware makes this possible. Based on our experience in developing the widely used macromolecular X-ray crystallography HKL data reduction package (Denzo, Scalepack, XdisplayF) we propose to further develop this area. The specific aims of this proposal are: 1) to develop data reduction methods for imperfect crystals - highly mosaic or twinned. it will be possible to index more than one diffraction pattern in one detector image, calculate overlap between diffraction patterns and correct for Bragg peak tails of reflection intruding upon each other. 2) to expand the Multiwavelength Anomalous Dispersion (MAD) method to cases where the signal-to-error ratio is currently insufficient for structure determination. This will be accomplished by calibrating and correcting for instrument response factors that now limit the MAD method to well diffracting crystals with relatively high anomalous diffraction signal. 3) to provide optimal data collection strategy by simulating in advance of data collection the expected reciprocal space coverage and detector resolution as a function of possible settings - data collection angles and crystal-to-detector distance. 4) to provide real-time feedback by reducing data on-line and calculating synthetic statistics. This is of particular significance to users of synchrotron facilities who need to wait for months to repeat their experiments. 5) to expand the range of the crystallographic experiments treated optimally in data reduction by deconvolution of partially overlapping reflections and 3-D profile fitting, to develop automatic and interactive editing tools in order to minimize the influence of possible experimental artifacts on the reduced data. 6) to expand methods to visually inspect the process of data collection and data reduction from current direct visualization of raw data to synthetic visual representation of factors affecting data quality - average reflection profiles, de convolution integrals, crystal slippage, detector response function and others. 7) to provide data base style organizational tools for the experimenter to follow the entire crystallographic project. The data base will be used to create unified reports - merging statistics, coverage, decay, slippage etc. Lastly, all of the above developments will be integrated into one efficient, simple to use computer and detector general software package. The resulting software will increase precision and reliability of three-D structures of macromolecules by x-ray crystallography, enable to solve crystal structures too difficult to solve with the present methods, and will speed-up, simplify and reduce the time and effort required of the structure determination process, speeding up understanding of biological systems and structure based drug design.